Fig. 3: SUMS identifies RgnTDC active site mutations that improve activity for a range of substrates.
From: Substrate multiplexed protein engineering facilitates promiscuous biocatalytic synthesis
a Active site model of RgnTDC (built from PDB ID: 4OBV)39 with residues highlighted at which mutations were found that significantly altered promiscuity or improved activity. b Select improved variants from active site libraries. Substrate screening conditions: 0.2 mM Trp and 7-I-Trp, and 2 mM 2-Me-Trp, 4-Br-Trp, 5-OMe-Trp, and 6-Cl-Trp, 4 h, 37 °C. Colored bars indicate the relative abundances of each product, and black diamonds indicate the total product formed. Full screening results found in Supplementary Figs. 6–S14. c Turnover numbers of wild-type RgnTDC and the top improved variant for each substrate. Different variants are depicted by different colored bars. Turnover numbers are presented as the averages of technical triplicate measurements with standard deviation shown as a bar. d Michaelis-Menten parameters for wild-type RgnTDC and activated variants for Trp and Trp analogs. Kinetic and turnover data were conducted in triplicate and complete data including error analysis are shown in Supplementary Tables 1 and 2 and Supplementary Figs. 17–21.
